For a bacteriophage to be useful for phage therapy it must be both isolated from the environment and shown to have certain characteristics beyond just killing strains of the target bacterial pathogen. These include desirable characteristics such as a relatively broad host range and a lack of other characteristics such as carrying toxin genes and the ability to form a lysogen. While phages are commonly isolated first and subsequently characterized, it is possible to alter isolation procedures to bias the isolation toward phages with desirable characteristics. Some of these variations are regularly used by some groups while others have only been shown in a few publications. In this review I will describe (1) isolation procedures and variations that are designed to isolate phages with broader host ranges, (2) characterization procedures used to show that a phage may have utility in phage therapy, including some of the limits of such characterization, and (3) results of a survey and discussion with phage researchers in industry and academia on the practice of characterization of phages.

Phages for Phage Therapy: Isolation, Characterization, and Host Range Breadth.

Gene cloning (DNA cloning) is nothing but the production of similar or exact copies (clones) of a particular gene or DNA sequence using genetic engineering techniques. In the genetic Engineering techniques the DNA containing the target gene is cut into fragments using restriction enzymes. With the help of the restriction enzymes these fragments are then inserted into cloning vectors, such as bacterial plasmids or bacteriophages, which transfer the recombinant DNA that is desirable gene to suitable host cells, such as the bacterium E. coli where the DNA gets replicated. Alternatively, the complementary DNA is inserted into the vectors, or ‘naked’ DNA fragments can be taken up directly by a host bacterium from its medium (this is less efficient than vector transfer).

A Review on Gene cloning and Genome Organization

Poultry and poultry products are the leading causes of foodborne salmonellosis worldwide. Antibiotics are used to control Salmonella spp. in poultry but its uncontrolled use results in the emergence of resistant pathogens.The use of bacteriophages as antimicrobial agents to control antibiotic resistant pathogenic bacteria could be a possible alternative. The aim of this study was to isolate, characterize and evaluate the effectiveness of bacteriophages for reducingload of Salmonella spp. on eggshells. One bacteriophage named as Sal-PE, specific to Salmonella enteritidiswas isolated from poultry excreta. For isolation, samples were subject to an enrichment protocol and then double agar layer method was performed to detect plaque. It had the capability to survive in wide range of pH between 4 to 10and found to be resistant at 60°C for 1 hour. Sal” PE showed its lytic effect on 13 of the 15 (87%) isolates including Salmonella enteritidis and Salmonella typhimurium which were recovered from 50 poultry excreta samples. After enrichment and growth on selective media, isolates were identified based on cultural characteristics, microscopic observation and biochemical tests. Amplification of three different genes (invA, sdfI, fliC) were carried out tocharacterize those isolates in molecular level. All isolates were found to be resistant to penicillin G, ampicillin, oxacillin and clindamycin but sensitive to ciprofloxacin, streptomycin, cefixime and chloramphenicol. Lytic efficiency of Sal-PE was determined by observing the reduction in optical density due to destruction of pathogens. Though more studies are needed in order to evaluate phage effectiveness, our findingsare expected to help us in initiating the development of a better preventive approach to control the occurrence of Salmonella spp. on eggshells. 
Bangladesh J Microbiol, Volume 35 Number 1 June 2018, pp 37-44

Isolation of Bacteriophage from Eggs and Use of Excreta as Biocontrol Agent in Controlling Eggs Related Salmonellosis

The genus Salmonella, as many others bacteria, has been frequently reported as resistant to commonly used antibiotics in poultry, therefore, the current spread of antibiotic resistance genes in bacteria, raises doubts on the effectiveness of antibiotics in the future. One possible alternative to antibiotics is the use of bacteriophages as antimicrobial agents to control antibiotic resistant pathogenic bacteria. The main goal of this project was to isolate and use bacteriophages as a new, safe, and effective biocontrol method for treating infectious diseases caused by Salmonella entero-pathogena. Four lytic bacteriophages cocktails (PSEA-2, SSEA, PSIA-2, SSIA) were isolated from wastewater of poultry processing industries, to control Salmonella enterica subsp enterica serovar Enteritidis (SE), and Salmonella enterica subsp enterica serovar Infantis (SI), under laboratory conditions. We found the cocktails PSEA-2 and PSIA-2 specific for SE and SI while cocktails SSEA and SSIA also caused lysis in Pseudomona. The observations in the Transmission Electron Microscope (TEM) revealed the presence of tailed phages of the Siphovididae and Myoviridae families; and a polyhedral phage. We have isolated specific phages and tested in vitro their effectiveness at controlling Salmonella. Further studies are needed in order to assess the phage effectiveness in vivo.

/pdf/isolation-of-lytic-bacteriophages-for-nanobiocontrol-of-4qn3gnhjki.pdf

Isolation of Lytic Bacteriophages for Nanobiocontrol of Pathogenic and Antibiotic Resistant Salmonella Present in Poultry in Ecuador

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Isolation of Lytic Bacteriophages for Nanobiocontrol of Pathogenic and Antibiotic Resistant Salmonella Present in Poultry in Ecuador